Nonalcoholic fatty liver disease (NAFLD) is a leading cause of chronic liver disease worldwide, yet its pathogenesis remains poorly understood. Impaired mitochondrial function is largely thought to be a core abnormality responsible for disease progression in this condition. The central question extant is what events link excessive lipid accumulation in liver cells to mitochondrial dysfunction. Thus, the overall objective of this proposal is to define the cellular and molecular mechanisms contributing to mitochondrial dysfunction and disease progression in NAFLD. Based on extensive preliminary data, we propose the novel CENTRAL HYPOTHESIS that excessive free fatty acids accumulation in the liver results in impaired mitochondrial function and NAFLD progression by triggering lysosomal permeabilization via regulation of the Bcl-2 family members We will now employ current and complementary, molecular, biochemical and cell biological approaches to further explore the lysosomal - mitochondrial axis in NAFLD. Our proposal has three SPECIFIC AIMS. FIRST, we will identify and manipulate novel intracellular targets that initiate lysosomal permeabilization in in-vitro and in-vivo models of NAFLD as well as human specimens of NAFLD and control individuals. SECOND, we will molecularly define the lysosomal - mitochondrial axis in models of NAFLD and cell free systems. FINALLY, we will determine if inhibition of lysosomal permeabilization and cathepsin B activation prevent liver injury and fibrosis in an in-vitro tissue model and in-vivo dietary murine models of NAFLD. The proposal is innovative technically and conceptually as it tests new concepts for lipid induced hepatotoxicity using sophisticated technologies. Moreover, because lipotoxicity as a result of over-accumulation of free fatty acids in non-adipose tissues has been implicated in the pathogenesis of other human liver diseases including chronic hepatitis C infection, alcoholic steatohepatitis and hemochromatosis, as well as other diseases such as type II diabetes and obesity associated heart disease the results of this proposal may not only bring new insights to the mechanisms underlying these conditions, but also could translate into new therapeutic strategies to treat them (e.g. the use of pharmacological Bax or cathepsin B inhibitors).